Transcriptomic Changes in ΔcomA and ΔrapA Mutants in Bacillus subtilis

Maintainer: Marek Gierlinski Collaborators: Jonathan Griffin, Nicola Stanley-Wall

Usage

The software in this repository is designed to be run in two stages. The first stage, including FASTQ quality control, cleaning, and mapping, is run on a computing cluster. The second stage, including all downstream analysis, is designed to be run in R on a local laptop or desktop.

On a Linux Cluster

Copy the contents of the rna_seq folder to the cluster filesystem. Change to the rna_seq directory and create and activate a conda environment:

cd rna_seq
conda create --name bsub_comp --file spec-file.txt
conda activate bsub_comp

Ensure that the FASTQ files are in the ./fastq subdirectory.

Run Snakemake:

./run_snake.sh

This will trim adapters, perform quality control, download genome files, map reads to the reference, and count reads per gene. Note: This shell script contains a call to snakemake with arguments configured for our Sun Grid Engine. If you're using a different cluster setup, you will need to modify the arguments accordingly.

On a Local Laptop/Desktop

Once Snakemake has finished, we recommend using Positron or RStudio for downstream analysis. If you’re working on a different machine (e.g., I run Positron on a laptop), you will need to copy some data using:

./scripts/get_data.sh

Note: Before using this script, edit it to change the remote location, as it currently points to my directory.

Once the mapping results are available on your local machine, open an R console in Positron or RStudio and create the environment using renv:

install.packages("renv")
renv::restore()

This will install all the required packages. If anything is missing, install additional packages locally using renv::install().

Once the environment is set up, run the targets pipeline:

targets::tar_make()

This will perform all calculations, generate data objects, figures, and tables (as targets objects), and output CSV files in the ./tab directory. An HTML report will be generated in the ./doc directory.